The storage and mobilization of lipid are fundamental cellular processes, and its dysregulation contributes to numerous diseases including diabetes, atherosclerosis and cardiomyopathy. Recent work indicates that the rate-limiting step for the mobilization of intracellular triglyceride is the release of Abhd5, a lipase co-activator, from specific lipid droplet scaffold proteins. We hypothesize that compounds that disrupt the association of Abhd5 with the scaffold proteins perilipin (Plin) and muscle lipid droplet protein (Mldp) will promote clearance of lipid from fat and muscle cells, respectively. We have developed a highly-sensitive assay based on protein complementation that can monitor the interaction of Abhd5 with Plin and Mldp in mammalian cells. This proposal seeks to adapt this assay to a cell-free system for high throughput screening. Active compounds will be characterized in secondary screens to demonstrate specificity, potency and efficacy. Compounds meeting criteria will be used as novel chemical probes to test specific mechanisms of cellular lipolysis in fat and muscle cells. Our long term goal is to identify lead compounds that will reduce lipid accumulation specifically in fat and muscle and thus would be potential novel treatments of obesity-related disorders including diabetes and cardiovascular disease.